J. clin. Path., 24, Suppl. (Roy. Coll. Path.), 5, 60-65

Absorption of cobalamins1. CHANARIN

From the Department of Haematology, Northwick Park Hospital and Clinical Research Centre, Harrow,Middlesex

Deoxyadenosylcobalamin (coenzyme B12) is thepredominant vitamin B12 analogue present in anormal diet, although small amounts of otheranalogues such as methylcobalamin and hydroxo-cobalamin are found (Weissbach, Toohey, andBarker, 1959; Toohey and Barker, 1961; Lindstrand,1964). Some hydroxocobalamin may arise as theresult of photolysis of the coenzyme forms. Thecobalamins are normally bound to protein in thecell (Cohn, Minot, Alles, and Salter, 1928; Hedbom,1960). These vitamin B12 analogues are absorbedlargely unchanged (Okuda, Yashima, and Taka-matsu, 1969), although some conversion of cyano-cobalamin to deoxyadenosylcobalamin during ab-sorption in the guinea pig has been demonstrated(Hoffbrand, Linnell, Matthews, and Peters, 1970).Further, the different vitamin B12 analogues areabsorbed equally by man (Lee and Glass, 1961;Chosy, Killander, and Schilling, 1962), althoughwhen assessed by urinary excretion methods differ-ences in plasma and tissue binding may result inlower urinary excretion with deoxyadenosyl B12 andhydroxocobalamin.An average 'low cost' mixed diet contained 16 ,ug

(range 1 2 to 75 6) per day (Chung, Pearson, Darby,Miller, and Goldsmith, 1961). Although a strictlyvegetarian diet may be almost devoid ofvitamin B12,some vitamin B12 may appear as the result of theincreasing bacterial content of the meal following itspreparation. Vegetarians may also get some vitaminB12 from water supplies, particularly water fromwells as in India. Cooking of food may serve torelease vitamin B12 from its protein link and there islittle loss of the vitamin during this process. Further,almost all the vitamin in food is available forabsorption. Thus vitamin B12 in meat (lamb) wasabsorbed to the same extent as the equivalent dosein aqueous solution (Heyssel, Bozian, Darby, andBell, 1966). As the daily vitamin B12 requirement isbetween 2 and 5 ,ug the amount present in an averagemixed diet is more than adequate and, indeed, theamount of vitamin B12 available often exceeds theabsorption capacity of the small gut.The amount of vitamin B12 that can be absorbed

60

from a single dose or a single meal by a physiologicalmechanism, that is, through the agency of intrinsicfactor, is about 2 ug. Thus with oral quantities ofvitamin B12 of less than 0 5 ug, over 70% is absorbedand this falls to 50% with 1 or 2 ug doses, 16% with10 ,ug, and less than 5% with doses of 20 Mug or more.With relatively large pharmacological doses ofvitamin B12 about 1 % may diffuse across the smallgut mucosa by a mechanism that does not requireintrinsic factor. The limitation to the amount ofvitamin B12 absorbed may be related to saturationof specific ileal receptor sites by intrinsic factorvitamin B12 complex. It has been suggested by Abels(1959) that the block to further vitamin B12 uptakelasts for some three hours.

Intrinsic Factor

The field has been reviewed by Chanarin (1968a,1968b, and 1969), Simons (1968), and Corcino,Waxman, and Herbert (1970). This glycoproteinwith a molecular weight of about 55,000 is secretedby the gastric parietal cell in man. Its secretion isaugmented by the polypeptide hormone gastrin andby its synthetic analogues such as pentagastrin, byhistamine and histalog, and by insulin. All thesesubstances are also potent stimulants of the othermajor product of the parietal cell, hydrochloric acid.Under these circumstances it is not surprising thatboth intrinsic factor and vitamin B12 are tolerant toa wide range ofpH and those changes in pH do notaffect the capacity of intrinsic factor to unite withvitamin B12 (McGuigan, 1967; Ashworth, Strick-land, Koo, and Taylor, 1969). However, an acid pHpromotes proteolytic digestion of intrinsic factor bypepsin in the gastric lumen and this results in a slowbut steady loss of intrinsic factor activity.The radioimmunoassay for intrinsic factor intro-

duced by Ardeman and Chanarin in 1963 has beenwidely employed in the diagnosis of perniciousanaemia and in the study of gastric secretion. A unitof intrinsic factor has been defined as the quantitybinding 1 nanogram of vitamin B12, and 400 to 500such units are required to promote optimum absorp-

copyright. on M

arch 12, 2021 by guest. Protected by

http://jcp.bmj.com

/J C

lin Pathol: first published as 10.1136/jcp.s3-5.1.60 on 1 January 1971. D

ownloaded from

Absorption of cobalamins

tion of 1 0 ,ug of vitamin B12 in pernicious anaemia(Ardeman and Chanarin, 1965a). On the other hand,the normal daily output of intrinsic factor is inexcess of 70,000 units with an average concentrationof 30 units per ml in the basal secretion and a con-centration of about 55 units per ml after a stimulusto secretion.

Interaction between Intrinsic Factor and Vitamin B12

The stoichiometric reaction between intrinsic factorand vitamin B12 which occurs with great rapidity invitro is largely complete in the stomach but can alsooccur in the small gut. Digestion in the stomachserves to release vitamin B12 in food, a process thatmay remain incomplete after partial gastrectomy.There may be some exchange between intrinsicfactor-bound vitamin B12 and free vitamin B12(Donaldson and Katz, 1963). A further importantaspect of the link between vitamin B12 and intrinsicfactor is that each becomes considerably more stable(Abels and Schilling, 1964). Thus, intrinsic factorwhen linked to vitamin B12 is much more resistantto proteolytic digestion as well as to heat, andadenosylcobalamin becomes resistant to photolysis(Okuda et al, 1969). It has also been suggested thatthe vitamin B12-intrinsic factor complex is less easilytaken up by bacteria than is free vitamin B12 (Boothand Heath, 1962). The complex passes down thesmall gut to be taken up by specific receptor sites onthe brush border of the epithelial cells of the villiin the ileum.

EVENTS IN THE ILEUMNormal intestinal uptake of the intrinsic factor-vitamin B12 complex depends not only on thepresence of the normal epithelial cells but on anormal milieu in the small gut lumen. Hyperacidity,as in the Zollinger-Ellison syndrome (Shimoda,Saunders, and Rubin, 1968), loss of pancreaticsecretion (Toskes and Deren, 1970), or interferencewith the integrity of the cell during colchicine therapy(Race, Paes, and Faloon, 1970) or by alcohol(Lindenbaum, Rybak, Gerson, Rubin, and Leiber,1970) all interfere with the ileal phase of vitamin B12absorption.The site of vitamin B12 absorption in man is the

distal ileum and the evidence has been reviewed byChanarin (1969). Following an oral dose of vitaminB12 detectable amounts of the vitamin appear in theblood some four hours later and reach a peak in eightto 12 hours (Booth and Mollin, 1956). This long lagis due to delay in passage of vitamin B12 from thesurface of the epithelial cell to the portal blood butthe precise events that occur are uncertain.

Specific receptors for the uptake of the vitamin3

B12-intrinsic factor complex appear to be present onthe microvillous membranes of the ileal epithelialcells. Thus, a homogenate of human ileum (but notjejunum) showed an enhanced uptake of intrinsicfactor-vitamin B12 that was inhibited by the additionof intrinsic factor antibody (Carmel, Rosenberg, Lau,Streiff, and Herbert, 1969). Donaldson, Mackenzie,and Trier (1967) localized this activity to the micro-villous fraction. Carmel et al (1969) found that cal-cium or magnesium ions were required for theattachment of the vitamin B12-intrinsic factor com-plex to human ileal homogenate and that the optimalpH was 6-6. Attempts to isolate an ileal receptorwere reported by Donaldson et al (1967) andby Rothenberg (1968).

It is believed that intrinsic factor separates fromvitamin B12 during the passage of vitamin B12 to theportal blood. This belief is based on repeated failureto demonstrate intrinsic factor in blood and the factthat some patients may have intrinsic factor antibodyin plasma while still having normal vitamin B12absorption (Ardeman, Chanarin, Krafchik, andSinger, 1965). Administration of hog intrinsic factorlabelled with 51Cr and vitamin B.2 labelled with57Co showed that vitamin B12 entered the blood butthat the chromium label was recovered from thefaeces (Yamaguchi, Rosenthal, and Glass, 1970).Peters and Hoffbrand (1970a) labelled human in-trinsic factor with 1251 and fed this with 57Co-labelled vitamin B12 to four guinea pigs. Fractiona-tion of the ileal cells appeared to show that the 1251label remained attached to the brush border whilethe 57Co label entered the cell and was attached tomitochondria. If confirmed, this would indicate thatseparation of intrinsic factor from vitamin B12 takesplace on the cell surface and that intrinsic factordoes not enter the epithelial cell. Further, since thisseparation preceded the lag in vitamin B12 transportto portal blood such separation may not be the causefor this lag. Vitamin B12 entering the epithelial cell isfound in the mitochondrial fraction (Peters andHoffbrand, 1970b). On the other hand if the modeof entry of vitamin B12 to the ileal epithelial cell wasby pinocytosis (Wilson, 1963) localization to lyso-somes would be anticipated, since pinocytoticvesicles fuse with lysosomes. Although the evidenceis against entry of vitamin B12 by pinocytosis, couldnot secondary localization of vitamin B12 to mito-chondria occur?Another hypothesis is that intrinsic factor is

separated by hydrolase enzymes on the brush border(Mackenzie and Donaldson, 1969). This is supportedby the preliminary observations of Peters andHoffbrand (1970a) that labelled intrinsic factorremains on the cell surface. Vitamin B12 entering theportal blood does so attached to a specific carrier

61

copyright. on M

arch 12, 2021 by guest. Protected by

http://jcp.bmj.com

/J C

lin Pathol: first published as 10.1136/jcp.s3-5.1.60 on 1 January 1971. D

ownloaded from

2L Chanarin

protein, transcobalamin II (Hall and Finkler, 1965).The transcobalamin II-vitamin B12 complex has avery short half-life so that absorbed vitamin B12 israpidly transferred to tissues such as the liver (Hall,1969; Hom, 1967). Transcobalamin II may besynthesized in the ileal epithelial cell and the complexwith vitamin B12 transferred to the blood. Lag inproduction of the carrier protein is another possibleexplanation of the long lag in delivery of absorbedvitamin B12 to blood.

Finally absorbed vitamin B12 reappears in the gutvia excretion in bile and this vitamin B12 is re-absorbed.

Vitamin B12 Malabsorption

This falls into three categories: (1) inadequateintrinsic factor which may be congenital, due togastric atrophy, gastric resection, or neutralizationby antibody; (2) luminal factors, such as abnormalsmall gut flora, fish tapeworm, pancreatitis, Zollinger-Ellison syndrome, laxatives (cascara), or drugs, eg,slow-K, EDTA; (3) ileal factors which can becongenital (Imerslund-Grasbeck), due to resection,gluten-sensitive enteropathy, tropical sprue, drugs,eg, ethanol, colchicine, para-aminosalicylic acid, orneomycin, or folate and vitamin B12 deficiency states.

INADEQUATE INTRINSIC FACTORFailure of intrinsic factor production may beinherited as a recessive characteristic and present asa familial form of megaloblastic anaemia in the firsttwo years of life. This has been reviewed by Chanarin(1969).

It has been recognized for some time that themalabsorption of vitamin B12 in pernicious anaemiais not fully corrected by additional intrinsic factor.There are a number of possible explanations for this.Free antibody to intrinsic factor in the gastro-intestinal tract has been found in a significant pro-portion of patients (Fisher, Rees, and Taylor, 1965;Schade, Feick, Muckerheide, and Schilling, 1966;Herbert, Carmel, and Li, 1967; Rose and Chanarin,1969; Goldberg and Bluestone, 1970). More recentlyRose and Chanarin (1971) have found a significantcorrelation between intrinsic factor antibody, particu-larly in gastric secretion, and the absorption ofvitamin B12 in pernicious anaemia. In the absence ofdemonstrable antibody the mean absorption ofvitamin B12 with intrinsic factor was the same as thatin healthy subjects. Absorption in the presence ofexcess intrinsic factor was most depressed in thosewith antibody in both serum and gastric juice. Thecorollary is the restoration of vitamin B12 absorptionin pernicious anaemia by steroid therapy and this isaccompanied by a steady decline in titre of intrinsic

factor antibody (Ardeman and Chanarin, 1965).Long-term vitamin B12 deficiency of itselfdepresses

the absorption of vitamin B12. This is correctedslowly over many months following the start ofvitamin B12 therapy and is a further factor con-tributing to impaired vitamin B12 absorption whengiven with intrinsic factor in pernicious anaemia(Haurani, Sherwood, and Goldstein, 1964; Brody,Estren, and Herbert, 1966; Carmel and Herbert,1967; Forshaw, 1969). A third contributory factor tointestinal malabsorption in pernicious anaemia maylie in the very high serum gastrin level in perniciousanaemia which of itself may produce intestinal mal-absorption (McGuigan and Trudeau, 1970; Wright,Hersh, Floch, and Weinstein, 1970).

Severe atrophic gastritis in the absence of per-nicious anaemia may lead to impaired intrinsic factorproduction (Ardeman and Chanarin, 1966) and halfthe patients without pernicious anaemia have mal-absorption of vitamin B12 (Chanarin, 1969). In asignificant number of these patients the absorptionof vitamin B12 is improved by an injection of car-bachol (Whiteside, Mollin, Coghill, Williams, andAnderson, 1964). However, carbachol does notstimulate intrinsic-factor secretion (Ardeman,Chanarin, and Doyle, 1964) butmay act by producingintestinal hurry and hence propelling small amountsof intrinsic factor-vitamin B12 complex to the ileum.

Megaloblastic anaemia due to vitamin B12deficiency develops in about 5% of patients under-going partial gastrectomy, although malabsorptionof vitamin B12 is present in some 30% of patientswhen the test is carried out in the fasting state (Lousand Schwartz, 1959). On the other hand absorptionof labelled vitamin B12 is improved when the dose isgiven with a meal (Deller, Germar, and Witts, 1961)or with an injection of histamine (Turnbull, 1967).This implies that a stimulus to intrinsic factor pro-duction may be necessary after partial gastrectomy.A further complication after partial gastrectomy isinterference with vitamin B12 absorption by anabnormal gut flora so that there is impaired absorp-tion of the vitamin B12-intrinsic factor complex. Thisis the case in 12% of patients who have impairedvitamin B12 absorption after partial gastrectomy(Chanarin, 1969).

LUMINAL FACTORSBoth infestation with the fish tape worm and anabnormal small-intestinal bacterial flora, by takingup dietary vitamin B12, make it unavailable to thehost. The disorders leading to an abnormal bacterialgut flora (reviewed by Chanarin, 1969) are associatedwith small gut stasis and the malabsorption may becorrected temporarily after administration of wide-spectrum antibiotics. These disorders are blind

62

copyright. on M

arch 12, 2021 by guest. Protected by

http://jcp.bmj.com

/J C

lin Pathol: first published as 10.1136/jcp.s3-5.1.60 on 1 January 1971. D

ownloaded from

Absorption of cobalamins

intestinal loops surgically produced or otherwise,strictures, entero-entero or entero-colic anastomosis,fistulae, small-intestinal diverticulosis, interference ofgut motility as in scleroderma, Whipple's disease,post-vagotomy, and after administration of ganglion-blocking agents.

Pancreatic insufficiency may be associated withimpaired vitamin B12 absorption (Maclntyre, Sachs,Krevans, and Conley, 1956; Perman, Gullberg,Reizenstein, Snellman, and AlIgen, 1960; Nieweg,Abels, Veeger, and Hillemans, 1962; Toskes andDeren, 1970). This was the case in nine out of 22consecutive patients with pancreatic insufficiencyreported by Toskes and Deren (1970) and in five outof nine patients studied by Perman et al (1960). Inthese cases the absorption of vitamin B12 is improvedby carrying out the absorption test with the additionof a single dose of pancreatic enzyme and also by theaddition of bicarbonate. Factors leading to impairedvitamin B12 absorption in pancreatitis may be anacid intestinal pH due to loss of bicarbonate secre-tion from the pancreas, and reduction in the amountof ionic calcium. However, Toskes and Deren (1970)have not found any difference in ileal pH and in totalcalcium concentration in those patients who ab-sorbed vitamin B12 normally and those who did not.They suggest that pancreatic extract contains a non-dialyzable, thermolabile factor needed for vitaminB12 absorption. Pancreatic enzymes conceivablycould be required for digestion of intrinsic factor-vitamin B12 complex as part of the process of thepassage of vitamin B12 into the ileal epithelial cell(LeBauer, Smith, and Greenberger, 1968).

Impaired absorption of vitamin B12 in theZollinger-Ellison syndrome was noted by Shimodaet al (1968) and was ascribed to the acid pH in thesmall gut. When the pH at the duodeno-jejunaljunction was maintained at 7 normal vitamin B12absorption was restored, the patient's own gastricsecretion serving as the source of intrinsic factor.

Administration of cascara was associated withincreased faecal excretion ofvitamin B12 although theurinary excretion remained within normal limits(Webb, Chodos, Mahar, and Faloon, 1968). If con-firmed, this would be a very unusual pattern butcould be explained by a failure to reabsorb vitaminB12 excreted via the bile.

ILEAL FACTORSA specific failure of transport of vitamin B12 acrossthe ileal cell (congenital vitamin B12 malabsorption)was described by Imerslund (1960) and by Grasbeck,Gordin, Kantero, and Kuhlback (1960). This is thecommonest cause of vitamin B12 malabsorption inchildren. It has a recessive mode of inheritance andthese children present with a severe megaloblastic

anaemia in the first or second year of life. Thestomach and gastric secretion is normal, that is, hasnormal content of acid and of intrinsic factor. Otherthan malabsorption of the intrinsic factor-vitaminB12 complex all other intestinal function tests arenormal. Proteinuria is a constant finding.

Ileal resection, for example for regional enteritisor following a volvulus, may lead to vitamin B12malabsorption. Involvement of the ileum in patientswith gluten-sensitive enteropathy results in impairedvitamin B12 absorption in about one third of patients.The majority of patients with tropical sprue haveimpaired vitamin B12 absorption and this may be theonly manifestation of the disease in the chronic state.There is improvement in the manifestations oftropical sprue with long-term antibiotic therapy(Klipstein, 1968), and although this evidence hasbeen cited as support for the view that an abnormalintestinal flora is the principal aetiological agent(Banwell and Gorbach, 1969) it is more probablethat an abnormal mucosa is responsible.A number of drugs or chemicals may lead to

vitamin B12 malabsorption. These include ethanol,slow-K, neomycin, colchicine, and ethylenediamine-tetraacetate (EDTA).

Depression of vitamin B12 absorption was foundin all five volunteers taking large amounts of alcohol(46 to 66% of total caloric intake) although nonewere inebriated (Lindenbaum et al, 1970). Abnormalvitamin B12 absorption was present in eight out of17 chronic alcoholics admitted to an Alcoholic Unit(Roggin, Iber, Kater, and Tabon, 1969). Ethanolprobably exerts a direct toxic effect on intestinal cellswhich, if an analogy can be drawn from its effect onthe marrow, is transient and reversible on ethanolwithdrawal.

Administration of colchicine also produces areversible malabsorption state, one of the mani-festations of which is malabsorption of vitamin B12.Thus 12 out of 13 subjects showed impaired absorp-tion reaching lowest values on the second to theseventh day after the start of oral colchicine (Webbet al, 1968). Normal vitamin B12 absorption wasrestored three to five days after withdrawal of thedrug. It is assumed that the effect is due to inter-ference with cell renewal on the ileal surface.Neomycin therapy produces intestinal malabsorp-

tion, one ofthe manifestations ofwhich is interferencewith vitamin B12 absorption (Jacobson, Chodos, andFaloon, 1960).

Since the original report by Heinivaara and Palva(1964) that para-aminosalicylic acid produced vita-min B12 malabsorption there have been contradictoryreports in the literature (Paaby and Norvin, 1966)but other evidence of malabsorption in suchpatients,including that of fat, was reported by Coltart (1969)

63

copyright. on M

arch 12, 2021 by guest. Protected by

http://jcp.bmj.com

/J C

lin Pathol: first published as 10.1136/jcp.s3-5.1.60 on 1 January 1971. D

ownloaded from

64 I. Chanarin

and Hess, Gregory, Thompson, and Welsh (1970).The effects were reversed on drug withdrawal andwere partly overcome by oral folic acid (Palva,Heinivaara, and Mattela, 1966; Hess et al, 1970).

Salokannel, Palva, and Takkunen (1970) reportedthat regular administration of potassium chloride(500 mg) in a slow release tablet led to impairedvitamin B12 absorption in 11 out of 30 patients inhospital. A fall in pH in the intestinal lumen wassuggested as a possible factor.Grasbeck and Nyberg (1958) found that ad-

ministration of EDTA when given orally withvitamin B12 interfered with its absorption possiblyby binding calcium ions.

Prolonged deficiency of either folate or vitaminB12 leads to impairment of the capacity of the ileumto absorb the vitamin B12-intrinsic factor complex.This is reversible after specific therapy but in manycases improvement becomes evident only after six to12 months of therapy. Lees (1961) noted impairedvitamin B12 absorption in epileptics which improvedafter folate therapy, and further examples wererecorded by Scott, Kammer, Burger, and Middleton(1968) in megaloblastic anaemia due to alcoholismand pregnancy and by Forshaw (1969) in nutritionalfolate deficiency.

Impaired uptake of the vitamin BL2-intrinsic factorcomplex by the ileum may result from vitamin B12deficiency due to fish tape worm infestation (Nyberg,Griisbeck, and Sippola, 1958), in pernicious anaemia(Brody et al, 1966; Lawrence, 1966; Forshaw, 1969),and in infants with congenital absence of intrinsicfactor (Lampkin and Maurer, 1967).

References

Abels, J. (1959). Intrinsic Factor van Castle en Resorptie van VitaminB1,. Thesis, Groningen.

Abels, J., and Schilling, R. F. (1964). Protection of intrinsic factor byvitamin B,,. J. Lab. clin. Med., 64, 375-384.

Ardeman, S., and Chanarin, I. (1963). A method for the assay ofhuman gastric intrinsic factor and for the detection andtitration of antibodies against intrinsic factor. Lancet, 2,1350-1354.

Ardeman, S., and Chanarin, I. (1965a). Assay of gastric intrinsicfactor in the diagnosis of Addisonian pernicious anaemia. Brit.J. Haemat., 11, 305-314.

Ardeman, S., and Chanarin, I. (1965b). Steroids and perniciousanemia. New Engi. J. Med., 273, 1352-1353.

Ardeman, S., and Chanarin, I. (1966). Intrinsic factor secretion ingastric atrophy. Gut, 7, 99-101.

Ardeman, S., Chanarin, I., and Doyle, J. C., (1964). Studies ongastric secretion of gastric intrinsic factor in man. Brit. med.J., 2, 600-603.

Ardeman, S., Chanarin, I., Krafchik, B., and Singer, W. (1965).Addisonian pernicious anaemia and intrinsic fa^tor antibodiesin thyroid disorders. Quart. J. Med., 35, NS, 421.431.

Ashworth, L. A. E., Strickland, R. G., Koo, N. C., and Taylor, K. B.(1969). Effect ofpH on intrinsic factor and non-intrinsic factorvitamin Bl, binding in human gastric juice neutralized in vivo.Gastroenterology, 57, 506-510.

Banwell, J. G., and Gorbach, S. L. (1969). Tropical sprue. Gut, 10,328-333.

Booth. C. C., and Heath, J. (1962). The effect of E. colt on theabsorption of vitamin B,,. Gut, 3, 70-73.

Booth, C. C., and Mollin, D. L. (1956). Plasma, tissue and urinaryradioactivity after oral administration of "Co-labelled vitaminB1,. Brit. J. Haemat., 2, 223-236.

Brody, E. A., Estren, S., and Herbert, V. (1966). Coexistent perniciousanemia and malabsorption in four patients: including onewhose malabsorption disappeared with vitamin B15 therapy.Ann. intern. Med., 64, 1246-1251.

Carmel, R., and Herbert, V. (1967). Correctable intestinal defect ofvitamin B1, absorption in pernicious anemia. Ann. intern. Med.,67, 1201-1207.

Carmel, R., Rosenberg, A. H., Lau, K.-S., Streiff, R. R., and Herbert,V. (1969). Vitamin B1, uptake by human small bowel homo-genate and its enhancement by intrinsic factor. Gastroenterology,56, 548-555.

Chanarin, I. (1968a). Intrinsic factor. Clin. chim. Acta, 22, 85-90.Chanarin, I. (1968b). Gastric intrinsic factor. Gut, 9, 373-375.Chanarin, I. (1969). The Megaloblastic Anaemias. Blackwell, Oxford.Chosy, J. J., Killander, A., and Schilling, R. F. (1962). Studies on

hydroxocobalamin. Estimates of absorption from the gut andbinding to serum and liver. In Vitamin B1, und intrinsic Faktor,Second European Symposium, Hamburg, 1961, edited by H. C.Heinrich, pp. 668-672. Enke, Stuttgart.

Chung, A. S. M., Pearson, W. N., Darby, W. J., Miller, 0. N., andGoldsmith, G. A. (1961). Folic acid; vitamin B,, pantothenicacid and vitamin B1, in human dietaries. Amer. J. clin. Nutr.,9, 573-582.

Cohn, E. J., Minot, G. R., Alles, G. A., and Salter, W. T. (1928). Thenature of the material in liver effeztive in pernicious anemia.II. J. biol. Chem., 77, 325-358.

Coltart, D. J. (1969). Malabsorption induced by para-aminosalicylate.Brit. med. J., 1, 825-826.

Corcino, J. J., Waxman, S., and Herbert, V. (1970). Absorption andmalabsorption of vitamin B1s. Amer. J. Med., 48, 562-569.

Deller, D. J., Germar, H., and Witts, L. J. (1961). Effect of food onabsorption of radioactive vitamin B1,. Lancet, 1, 574-577.

Donaldson, R. M., and Katz, J. H. (1963). Exchange between freeand gastric juice-bound cyanocobalamin. J. clin. Invest., 42,534-545.

Donaldson, R. M., Jr., Mackenzie, I. L., and Trier, J. S. (1967).Intrinsic factor mediated attachment of vitamin B1, to brushborders and microvillous membranes of hamster intestine. J.clin. Invest., 46, 1215-1228.

Fisher, J. M., Rees, C., and Taylor, K. B. (1965). Antibodies ingastric juice. Science, 150, 1467-1469.

Forshaw, J. (1969). Effect of vitamin B1, and folic acid deficiency onsmall intestinal absorption. J. clin. Path., 22, 551-553.

Goldberg, L. S., and Bluestone, R. (1970). Hidden gastric auto-antibodies to intrinsic factor in pernicious anemia. J. Lab. clin.Med., 75, 449-456.

Grasbeck, R., Gordin, R., Kantero, I., and Kuhlback, B. (1960).Selective vitamin B1, malabsorption and proteinuria in youngpeople. A syndrome. Acta med. scand., 167, 289-296.

Grasbeck, R., and Nyberg, W. (1958). Inhibition of radiovitamin B1,absorption by ethylenediaminetetraacetate (EDTA) and itsreversal by cal-ium ions. Scand. J. clin. Lab. Invest., 10, 448.

Hall, C. A. (1969). Transport of vitamin B1, in man. Brit. J. Haemat.,16, 429-433.

Hall, C. A., and Finkler, A. E. (1965). The dynamics oftranscobalaminII. A vitamin B1, binding substance in plasma. J. Lab. clin.Med., 65, 459468.

Haurani, F. I., Sherwood, W., and Goldstein, F. (1964). Intestinalmalabsorption ofvitamin B1, in pernicious anemia. Metabolism,13, 1342-1348.

Hedbom, A. (1960). A native cobalamin-polypeptide complex fromliver: isolation and characterization. Biochem. J., 74, 307-312.

Heinivaara, O., and Palva, I. P. (1964). Malabsorption of vitamin B,,during treatment with para-aminosalicylic acid. A preliminaryreport. Acta med. scand., 175, 469-471.

Herbert, V., Carmel, R., and Li, L. G. (1967). Antibody causingvitamin B1, deficiency. New Engl. J. Med., 276, 61.

Hess, D. R., Gregory, D. M., Thompson, J. B., and Welsh, J. D.(1970). Para-aminosalicylic acid induced intestinal mal-absorption. Clin. Res., 18, 77.

Heyssel, R. M., Bozian, R. C., Darby, W. J., and Bell, M. C. (1966).Vitamin B12 turnover in man. The assimilation of vitamin B1,from natural foodstuff by man and estimates of minimal dailydietary requirements. Amer. J. clin. Nutr., 18, 176-184.

Hoffbrand, A. V., Linnell, J. C., Matthews, D. M., and Peters, T. J.(1970). The forms of vitamin B1, in the ileal enterocyte of the

copyright. on M

arch 12, 2021 by guest. Protected by

http://jcp.bmj.com

/J C

lin Pathol: first published as 10.1136/jcp.s3-5.1.60 on 1 January 1971. D

ownloaded from

Absorption of cobalamins 65

guinea pig during fasting and absorption of cyanocobalamin.J. Physiol. (Lond.), 208, 66P-67P.

Hom, B. L. (1967). Plasma turnover of 1'Cobalt-vitamin B,, boundto transcobalamin I and II. Scand. J. Haemat., 4, 321-332.

Imerslund, 0. (1960). Idiopathic chronic megaloblastic anemia inchildren. Acta paediat. scand., 49, Suppl. 119, 1-115.

Jacobson, E. D., Chodos, R. B., and Faloon, W. W. (1960). Anexperimental malabsorption syndrome induced by neomycin.Amer. J. Med., 28, 524-533.

Klipstein, F. A. (1968). Tropical sprue. Gastroenterology, 54, 275-293.Lampkin, B. C., and Maurer, A. M. (1967). Congenital pernicions

anemia with coexistent transitory intestinal malabsorption ofvitamin B12. Blood, 30,495-502.

Lawrence, C. (1966). B1, binding protein deficiency in perniciousanemia. Blood, 27, 389-394.

LeBauer, E., Smith, K., and Greenberger, N. J. (1968). Pancreaticinsufficiency and vitamin B,, malabsorption. Arch. intern. Med.,122, 423-425.

Lee, D. H., and Glass, G. B. J. (1961). Hepatic uptake and intestinalabsorption of co"-labelled 5,6-dimethylbenzimidazolylcoba-mide coenzyme. Proc. Soc. exp. Biol. (N. Y.), 107, 293-296.

Lees, F. (1961). Radioactive vitamin B1, absorption in the megalo-blastic anaemia caused by anticonvulsant drugs. Quart. J. Med.,30, 231-243.

Lindenbaum, J., Rybak, B., Gerson, C. D., Rubin, E., and Lieber,C. S. (1970). Effects of ethanol on the small intestine of man.Clin. Res., 18, 385.

Lindstrand, K. (1964). Isolation of methylcobalamin from naturalsource material. Nature (Lond.), 204, 188-189.

Lous, P., and Schwartz, M. (1959). The absorption of vitamin B1,following partial gastrectomy. Acta med. scand., 164, 407417.

Maclntyre, P. A., Sachs, M. V., Krevans, J. R., and Conley, C. L.(1956). Pathogenesis and treatment of macrocytic anemia.Arch. intern. Med., 98, 541-549.

Mackenzie, I. L., and Donaldson, R. M. (1969). Vitamin B,, absorp-tion and the intestinal cell surface. Fed. Proc., 28, 4145.

McGuigan, J. E. (1967). Measurement of the affinity of human gastricintrinsic factor for cyanocobalamin. J. Lab. clin. Med., 70,666-672.

McGuigan, J. E., and Trudeau, W. L. (1970). Serum gastrin con-centrations in pernicious anemia. New Engl. J. Med., 282,358-361.

Nieweg, H. O., Abels, J., Veeger, W., and Hellemans, N. (1962). Thedefective absorption of vitamin B,, in pancreatic insufficiency.In Vitamin B1, und intrinsic Faktor, Second European Sym-posium, Hamburg, 1961, edited by H. C. Heinrich, pp. 610-612.Enke, Stuttgart.

Nyberg, W., Grasbeck, R., and Sippola, V. (1958). Urinary excretionof radiovitamin B12 in carriers of diphyllobothrium latum. NewEngl. J. Med., 259, 216-219.

Okuda, K., Yashima, K., and Takamatsu, M. (1969). Intestinalabsorption of cobamnide coenzyme in relation to structuralintegrity and intrinsic factor. J. Lab. clin. Med., 74, 218-228.

Paaby, P., and Norvin, E. (1966). The absorption of vitamin B,,during treatment with para-aminosalicylic acid. Acta med.scand., 180, 561-564.

Palva, I. P., Heinivaara, O., and Mattela, M. (1966). Drug-inducedmalabsorption of vitamin B1,. III. Interference of PAS andfolic acid in the absorption of vitamin B12. Scand. J. Haemat.,3, 149-153.

Perman, G., Gullberg, R., Reizenstein, P. G., Snellman, B., andAlIgen, L.-G. (1960). A study of absorption patterns in mal-absorption syndromes. Acta med. scand., 168, 117-125.

Peters, T. J., and Hoffbrand, V. (1970a). Absorption of vitamin B,,in the guinea pig. Role of the mitochondrion. In Proceedingsof Glaxo Symposium. Churchill, London. (In press.)

Peters, T. J., and Hoffbrand, V. (1970b). Absorption of vitamin B1,by the guinea pig. I. Subcellular localization of vitamin B1, inthe ileal enterocyte during absorption. Brit. J. Haemat., 19,369-382.

Race, T. F., Paes, I. C., and Faloon, W. W. (1970). Intestinal mal-absorption induced by oral colchicine. Comparison withneomycin and cathartic agents. Amer. J. med. Sci., 259, 32-41.

Roggin, G. M., Iber, F. L., Kater, R. M. H., and Tabon, F. (1969).Malabsorption in the chronic alcoholic. Johns Hopk. med. J.,125, 321-330.

Rose, M. S., and Chanarin, I. (1969). Dissociation of intrinsic factorfrom its antibody: application to study of pernicious anaemiagastric juice specimens. Brit. med. J., 1, 468-470.

Rose, M. S., and Chanarin, I. (1971). The effect of intrinsic factorantibody on vitamin B1, absorption in pernicious anaemia.Brit. med. J., 1, 25-26.

Rothenberg, S. P. (1968). Identification of a macromolecular factorin the ileum which binds intrinsic factor and immunologicidentification of intrinsic factor in ileal extracts. J. clin. Invest.,47, 913-923.

Salokannel, S. J., Palva, I. P., and Takkunen, J. T. (1970). Mal-absorption of vitamin B1, during treatment with slow-releasepotassium chloride. Acta med. scand., 187, 431-432.

Schade, S. G., Feick, P., Muckerheide, M., and Schilling, R. F. (1966).Occurrence in gastric juice of antibody to a complex of in-trinsic factor and vitamin B12. New Engl. J. Med., 275, 528-531.

Scott, R. B., Kammer, R. B., Burger, W. F., and Middleton, F. G.(1968). Reduced absorption of vitamin B1, in two patients withfolic acid deficiency. Ann. intern. Med., 69, 111-114.

Shimoda, S. S., Saunders, D. R., and Rubin, C. E. (1968). TheZollinger-Ellison syndrome with steatorrhoea. II. The mechan-isms of fat and vitamin B12 malabsorption. Gastroenterology,55, 705-723.

Simons, K. (1968). Gastric intrinsic factor and other vitamin B,transport proteins: Chemical and physiological properties. InProgress in Gastroenterology, edited by G. J. Glass. Grune andStratton, New York.

Toohey, J. I., and Barker, H. A. (1961). Isolation of coenzyme B1,from liver. J. biol. Chem., 236, 560-563.

Toskes, P. P., and Deren, J. J. (1970). The role of the pancreas invitamin B1, absorption. Clin. Res., 18, 391.

Turnbull, A. L. (1967). Absorption of vitamin B1, in patients withanaemia after polya partial gastrectomy. Brit. J. Haemat., 13,752-763.

Webb, D. I., Chodos, R. B., Mahar, C. Q., and Faloon, W. W. (1968).Mechanism of vitamin B1, malabsorption in patients receivingcolchicine. New Engl. J. Med., 279, 845-850.

Weissbach, H., Toohey, J. I., and Barker, H. A. (1959). Isolation andproperties of B12 coenzymes containing benzimidazole anddimethylbenzimidazole. Proc. nat. Acad. Sci. (Wash.), 45,521-525.

Whiteside, M. G., Mollin, D. L., Coghill, N. F., Williams, A. W.,and Anderson, B. (1964). The absorption of radioactivevitamin B1, and the secretion of hydrochloric acid in patientswith atrophic gastritis. Gut, 5, 385-399.

Wilson, T. H. (1963). Intestinal absorption of vitamin B1,. Physio-logist, 6, 11-26.

Wright, H. K., Hersh, T., Floch, M. H., and Weinstein, L. D. (1970).Impaired intestinal absorption in the Zollinger-Ellison syn-drome independent of gastric hypersecretion. Amer. J. Surg.,119, 250-253.

Yamaguchi, N., Rosenthal, W. S., and Glass, G. B. J. (1970). Studyof the intestinal absorption of "'Cr-labeled intrinsic factor.Amer. J. clin. Nutr., 23, 156-164.

copyright. on M

arch 12, 2021 by guest. Protected by

http://jcp.bmj.com

/J C

lin Pathol: first published as 10.1136/jcp.s3-5.1.60 on 1 January 1971. D

ownloaded from